Thrombopoietin (TPO) is a factor that enhances the differentiation and maturation of megakaryocytes (platelet precursor cells) from hemopoietic stem cells into platelets. TPO also functions as a cytokine with an important role in the regulation of platelet number. TPO is converted into its active form through the cleavage of a TPO precursor comprising 353 amino acids.
Mpl is a TPO receptor, and human Mpl molecules are known to exist in two forms comprising 572 and 635 amino acids. The human Mpl gene sequence has already been analyzed (see Non-Patent Document 1 and GenBank accession No. NM—005373).
Most cytokine receptors dimerize upon ligand binding, and transduce signals into cells. It has been reported that TPO similarly binds to its own specific receptor MPL, which leads to dimerization of the receptor, thereby transducing signals into cells and exerting physiological effects (see Non-Patent Document 2).
Antibodies exhibiting agonistic activity have been reported among those antibodies that bind to receptors having the above features.
For example, an antibody against the erythropoietin (EPO) receptor has been reported to substitute for erythropoietin function. The monovalent form (Fab) of the antibody is capable of binding to the EPO receptor but is unable to transduce signals. Thus, dimerization of the erythropoietin receptor via bivalent binding is assumed to be essential for signal transduction (see Non-Patent Document 3).
Antibodies that bind to Mpl and exhibit TPO agonistic activity have also been reported (see Non-Patent Documents 4 and 5). This suggests that receptor dimerization is induced upon binding of a bivalent antibody with regards to MPL as well.
Meanwhile, a single-chain antibody (scFv) has been reported to exhibit TPO agonistic activity (see Patent Document 1). However, it has been revealed that, the underlying mechanism of scFv exhibiting TPO agonistic activity is that a part of scFv dimerizes (diabody) and this diabody becomes the actual active unit (see Patent Documents 2 to 4).    [Patent Document 1] U.S. Pat. No. 6,342,220    [Patent Document 2] WO 01/79494    [Patent Document 3] WO 02/33072    [Patent Document 4] WO 02/33073    [Non-Patent Document 1] Palacios et al., Cell, 1985, 41, 727-734    [Non-Patent Document 2] Souyri et al., Cell, 1990, 63, 1137-1147    [Non-Patent Document 3] Elliott, S. et al., J. Biol. Chem., 1996, 271(40), 24691-24697    [Non-Patent Document 4] Abe et al., Immunol. Lett., 1998, 61, 73-78    [Non-Patent Document 5] Bijia Deng et al., Blood, 1998, 92, 1981-1988